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Li, Ling, Sun, Di, Cong, Yu, and Cong: Characterization of Newcastle disease virus obtained from toco toucan
Original Article

J Vet Sci 2020;21(2):e19.

Published online: 4 March 2020

DOI: https://doi.org/10.4142/jvs.2020.21.e19

Characterization of Newcastle disease virus obtained from toco toucan

Jiaxin Li1, †, Mengmeng Ling1, †, Yixue Sun2, †, Haiyang Di3, Yulin Cong1, Haiying Yu1, Yanlong Cong1, *

1Laboratory of Infectious Diseases, College of Veterinary Medicine, Key Laboratory of Zoonosis Research, Ministry of Education, Jilin University, Changchun 130062, China

2Jilin Academy of Animal Husbandry and Veterinary Medicine, Changchun 130062, China

3Zoological and Botanical Garden of Changchun, Changchun 130062, China

*Corresponding author: Yanlong Cong Laboratory of Infectious Diseases, College of Veterinary Medicine, Key Laboratory of Zoonosis Research, Ministry of Education, Jilin University, 5333 Xi'an Road, Lvyuan, Changchun 130062, China. E-mail: congyanlong@126.com

† Jiaxin Li, Mengmeng Ling, and Yixue Sun contributed equally to this work.

Received 16 October 2019       Revised 6 December 2019       Accepted 16 December 2019

Copyright © 2020 The Korean Society of Veterinary Science

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Given that the current Newcastle disease virus (NDV) infection in wild birds poses the threat to poultry, surveillance of Newcastle disease in captive wild birds was carried out in Jilin, China in 2018. Here, an NDV strain obtained from toco toucan was firstly characterized. The results showed that the F gene of the NDV isolate Toucan/China/3/2018 is classified as genotype II in class II. Sequence analysis of the F0 cleavage site was 113 RQGR/L117, which supports the result of the intracerebral pathogenicity index assay indicating classification of the isolate as low-pathogenicity. Experimental infection demonstrated that Toucan/ China/3/2018 can effectively replicate and transmit among chickens. To our knowledge, this is the first report on genetically and pathogenically characterizing NDV strain isolated from toucan, which enriches the epidemiological information of NDV in wild birds.

Keywords: Toucan, NDV, genetic analysis, transmission potential

References

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Fig. 1.
Phylogenetic analysis based on the open reading frame (1–1,662 nt) of F genes from 20 genotypes of Newcastle disease virus. The tree was created by the maximum-likelihood method and bootstrapped with 1,000 replicates. Toucan isolate analyzed in the present study is in italics.
jvs-21-e19f1.tif
Fig. 2.
Phylogenetic analysis based on the full-length (1–1,662 nt) sequence of F genes from genotype II of Newcastle disease virus available in GenBank. The tree was created by the maximum-likelihood method and bootstrapped with 1,000 replicates. Only bootstrap values above 50 are shown. The vaccine strains are shown as solid black circles. Toucan strain analyzed in the present study is shown as a solid black triangle.
jvs-21-e19f2.tif
Fig. 3.
Immunohistochemistry analysis of tissue samples from chickens intranasally inoculated with Toucan/China/3/2018 at 7 days post inoculation. Arrows indicate the positive labeling of Newcastle disease virus. (A) Brain; (B) thymus; (C) trachea; (D) lung; (E) heart; (F) liver; (G) spleen; (H) glandular stomach; (I) small intestine; (J) pancreas; (K) kidney; and (L) bursa of Fabricius. Arrows indicate magnification, 20×.
jvs-21-e19f3.tif
Table 1.
Primers used for amplifying the Newcastle disease virus strain of Toucan/China/3/2018 in the study
Name Primer sequence (5′-3′) Genomic location
1F ACCAAACAGAGAATCCG 1–17
1R GTTGTTGGTGAGCCGC 1,757–1,772
2F CACATGACCACACCCTC 1,666–1,683
2R GATTAATTACAGTTGAGCGATC 3,206–3,228
3F GACCTAAGGTCCAACTC 3,148–3,164
3R CTAACTGTAAAGAATCAGG 4,451–4,469
4F TGCGTCTCTGAGATTGCG 4,387–4,404
4R CGACTGAAGGTAGAGTTACC 5,401–5,420
5F CAGCAGCGGCTTAATCAC 5,335–5,352
5R AGCTGACAGACTACCAGAG 6,253–6,271
6F CACAGATGAGGAACGAAGG 6,207–6,225
6R CTGAGAACCATACGCGG 7,341–7,357
7F GGTTGTGATATGCTGTGCTC 7,147–7,166
7R GCCTTGTATCTCATTGCCAC 8,328–8,347
8F GTTGCCAGTTGACCACAATC 8,245–8,264
8R CTTAGCGAAGATCCGTCC 10,031–10,048
9F GACGACCCTTGAGTACCTTAG 9,955–9,975
9R CGTGCATAGTCTGCCAGTG 11,724–11,742
10F TGCGGATAGTCAATTATTCTAG 11,616–11,637
10R GCACCAATATCTTGCACAG 13,437–13,456
11F GCTAATCTGTATTACATGTC 13,325–13,344
11R ACCAAACAAAGATTTGGTG 15,168–15,186
Table 2.
Genomic characteristics of Toucan/China/3/2018
Regions Gene sequence (nt) 3′ UTR (nt) ORF (nt) 5′ UTR (nt) Intergenic region (nt) Nucleotide length (nt) Amino acid length (aa)
Leader 1–55 – – – – 55 –
NP 56–1,801 66 122–1,591 210 2 1,746 489
P 1,804–3,254 83 1,887–3,074 180 1 1,451 395
M 3,256–4,496 34 3,290–4,384 112 1 1,241 364
F 4,498–6,289 46 4,544–6,205 84 31 1,792 553
HN 6,321–8,322 91 6,412–8,130 192 47 2,002 572
L 8,370–15,072 11 8,381–14,995 77 – 6,703 2,204
Trailer 15,073–15,186 – – – – 114 –

UTR, untranslated region; ORF, open reading frame.

Table 3.
Frequency of NDV isolation and virus titers from chickens challenged and those for cohabitation infection
Dpi Chickens
Intranasal inoculation Cohabitation infection
O C O C
Frequency of NDV isolation HA titers (nlog2) Frequency of NDV isolation HA titers (nlog2) Frequency of NDV isolation HA titers (nlog2) Frequency of NDV isolation HA titers (nlog2)
1 0/6 – 0/6 – 0/6 – 0/6 –
2 1/6 1.0 0/6 – 0/6 – 0/6 –
3 1/6 1.5 1/6 2.0 0/6 – 0/6 –
4 3/6 2.0 3/6 2.0 1/6 1.5 0/6 –
5 4/6 2.5 5/6 2.0 2/6 2.0 1/6 2.0
6 5/6 4.0 6/6 3.0 4/6 2.0 3/6 2.0
7 5/6 3.5 6/6 2.5 3/6 2.5 5/6 3.0
8 4/6 3.0 6/6 3.0 3/6 2.0 6/6 2.5
10 2/6 2.0 5/6 2.5 2/6 2.0 4/6 3.0
12 1/6 2.5 3/6 3.0 1/6 2.0 3/6 2.5
14 1/6 2.0 2/6 2.5 0/6 – 2/6 3.0

NDV, Newcastle disease virus; dpi, days post inoculation; O, oropharyngeal swab; C, cloacal swab; -, no hemagglutination activity.

Table 4.
Comparison of amino acid substitutions in the functional domains of the F protein
Strains HRc (471–500 aa) Major transmembrane domain (501–521 aa)
486 520
LaSota R I
Toucan/China/3/2018 S V

HRc, heptad repeat C region.

Table 5.
Comparison of amino acid substitutions in the functional domains of the HN protein
Strains Antigenic sites
1 2 3 4 12 14 23
345 513 514 521 569 263 287 321 332 333 356 494 516 347 350 353 193 194 201
LaSota P R I S D N D K G K K G R E Y R L S H
Toucan/China/3/2018 – – – – – – – – – – – – – – – – – – –

Dots indicate residues identical to LaSota.

Table 6.
Cross-HI test and coefficients of antigenic relatedness (R%) between Toucan/China/3/2018 and vaccine strain LaSota
Viruses Antisera Coefficient of antigenic relatedness
Toucan/China/3/2018 LaSota
Toucan/China/3/2018 10,240 10,240 70.71%
LaSota 1,280 2,560 70.71%
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